23 research outputs found
Identifying a Structural Preference in Reduced Rare-Earth Metal Halides by Combining Experimental and Computational Techniques
Stability measurements of antisense oligonucleotides by capillary gel electrophoresis.
The approach of using antisense oligonucleotides as potential drugs is based on hybridization of a short chemically-modified oligonucleotide with complementary cellular DNA or RNA sequences. A critical question is the stability of chemically modified antisense oligonucleotides in cellular environments. In a model system, resistance against various nucleases was evaluated by capillary gel electrophoresis (CGE). For some of the samples, matrix assisted laser desorption and ionization mass spectrometry (MALDI-MS) was used as an additional analytical tool to perform stability measurements. Using CGE, the enzymatic degradation of single nucleotides from the oligomer can be followed after different incubation times. 10% T polyacrylamide gels give baseline resolution for oligonucleotides ranging between 5 and 30 bases in length. The kinetic influence of a specific nuclease concentration and the antisense oligonucleotide structure on the cleavage reaction are discussed. Also, a simple desalting method to improve the injection efficiency and sensitivity of the method are described. Examples of measurements of chemically modified antisense 19-mers are presented
Influence of solvents and detergents on matrix-assisted laser desorption/ionization mass spectrometry measurements of proteins and oligonucleotides.
The effect of solvents was found to be critical for sample preparation in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). For proteins and oligonucleotides the use of 2-propanol/water as a solvent for different matrices can significantly improve the quality of spectra. This effect is demonstrated with proteins ranging in molecular weight from 12 to 150 kDa and with a special 19-mer oligonucleotide. A comparison of MALDI-MS using of 2-propanol as matrix solvent and high-performance capillary electrophoresis resulted in identical relative peak intensities for a p(dT)12-18 oligonucleotide mixture. Additionally, the effect of detergents for characterization of high molecular weight proteins in very dilute solutions was studied with this solvent. It was found that Triton X-100, up to a concentration of 1%, was highly compatible with MALDI measurements and even could improve the quality of spectra. Use of detergents for cell profiling has extended the detectable mass range to about m/z 75,000